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Intra-Bone Marrow Transplantation of Endosteal Bone Marrow Cells Facilitates Allogeneic Hematopoietic and Stromal Cells Engraftment Dependent on Early Expression of CXCL-12.

Chen C, Su Y, Chen J, Zhang D, Song Y, Guo S - Med. Sci. Monit. (2015)

Bottom Line: We compared the new method with each control group for allogeneic HSCs homing pattern, peripheral blood chimerism level, skin allograft survival time, and donor stromal cell percentage in recipient BM.By AMD3100 blockade at day 1, peripheral blood chimerism level and donor stromal cell percentage were significantly reduced as compared to the control group without AMD3100 blockade.Our study suggests that IBBMT of endosteal BMCs is an effective approach for HSCT in inducing allogeneic hematopoietic reconstitution.

View Article: PubMed Central - PubMed

Affiliation: Department of Plastic and Reconstructive Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, China (mainland).

ABSTRACT

Background: Hematopoietic stem cell transplantation (HSCT) has been considered as an effective approach at inducing allogeneic hematopoietic reconstitution and immune tolerance. However, it remains critical to find the optimal HSCT delivery method and robust sources of hematopoietic stem cells (HSCs).

Material and methods: We introduced a new method by infusing allogeneic endosteal bone marrow cells (BMCs) harvested from long bones endosteum through intra-bone marrow transplantation (IBBMT) into irradiated mice. Recipient mice that were transplanted with central BMCs or through intravenous bone marrow transplantation (IVBMT) were used as controls (n=6 per group). We compared the new method with each control group for allogeneic HSCs homing pattern, peripheral blood chimerism level, skin allograft survival time, and donor stromal cell percentage in recipient BM. AMD3100 was injected to determine whether chemokine stromal cell-derived factor-1 (CXCL-12) was critical for the new method.

Results: More allogeneic HSCs homed into spleen and bone marrow for the new method as compared to each control group. IBBMT of endosteal BMCs led to a higher peripheral blood chimerism and skin allograft survival. At 18 weeks, donor stromal cell percentage in recipient BMCs was higher for the new method than in each control group. By AMD3100 blockade at day 1, peripheral blood chimerism level and donor stromal cell percentage were significantly reduced as compared to the control group without AMD3100 blockade.

Conclusions: Our study suggests that IBBMT of endosteal BMCs is an effective approach for HSCT in inducing allogeneic hematopoietic reconstitution. The advantage is dependent upon the early expression of CXCL-12 after bone marrow transplantation.

No MeSH data available.


Related in: MedlinePlus

The analysis of recipient stromal cell replacement and gene expressions of recipient BMCs. (A) The gating protocol for donor-derived stromal cells in recipient BMCs. FSC, Forward Scatter. SSC, Side Scatter. Blue line, isotype control. (B, C) Donor stromal percentage in recipient BMCs were assessed at 1 (B) and 18 (C) weeks after BMT. Donor stromal cell population were H-2kb+ CD45-CD31-TER119- cells in BMCs harvested from the left tibia BMs. *, p value for IB-eBMCs versus each control group, p<0.05. Each dot represents an individual mouse result and bars are the mean for each group (%), n=6 per group. (D–G) Quantitative PCR analyzes of CXCL-12 (D, E) and n-cadherin (F, G) expression of recipient BMCs were performed at 1 week (D, F) and 18 weeks (E, G) after BMT. The corresponding gene expression level of BMCs from normal untreated mice was taken as control after normalization to GAPDH. All the bars represent the mean folds increase relative to that of normal mice BMCs for each group, n=6–8 per group. *, p value of IB-eBMCs group versus each control group (IB-cBMCs, IV-eBMCs, IV-cBMCs group respectively), p<0.05. ns is for not significant.
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f1-medscimonit-21-2757: The analysis of recipient stromal cell replacement and gene expressions of recipient BMCs. (A) The gating protocol for donor-derived stromal cells in recipient BMCs. FSC, Forward Scatter. SSC, Side Scatter. Blue line, isotype control. (B, C) Donor stromal percentage in recipient BMCs were assessed at 1 (B) and 18 (C) weeks after BMT. Donor stromal cell population were H-2kb+ CD45-CD31-TER119- cells in BMCs harvested from the left tibia BMs. *, p value for IB-eBMCs versus each control group, p<0.05. Each dot represents an individual mouse result and bars are the mean for each group (%), n=6 per group. (D–G) Quantitative PCR analyzes of CXCL-12 (D, E) and n-cadherin (F, G) expression of recipient BMCs were performed at 1 week (D, F) and 18 weeks (E, G) after BMT. The corresponding gene expression level of BMCs from normal untreated mice was taken as control after normalization to GAPDH. All the bars represent the mean folds increase relative to that of normal mice BMCs for each group, n=6–8 per group. *, p value of IB-eBMCs group versus each control group (IB-cBMCs, IV-eBMCs, IV-cBMCs group respectively), p<0.05. ns is for not significant.

Mentions: All staining was performed at a cell concentration of 1×106/ml. All peripheral blood marrow cells (PBMCs) were gated according to the protocol described by Krause [31]. The peripheral blood chimerism level was calculated as H-2kb + cells percentage (%) in all PBMCs. The donor HSCs phenotype was H-2kb+ c-kit+ Sca-1+ Lineage- BMCs. Donor CFSE-positive cells were calculated as CFSE-positive cells in CD45+ cell population. The gate was set according to isotype control. The flow cytometry gating protocol for donor stromal cells in recipient BMCs is shown in Figure 1A. Donor stromal cell population are represented as H-2kb+CD45-CD31-TER119- cells in all BMCs (%).


Intra-Bone Marrow Transplantation of Endosteal Bone Marrow Cells Facilitates Allogeneic Hematopoietic and Stromal Cells Engraftment Dependent on Early Expression of CXCL-12.

Chen C, Su Y, Chen J, Zhang D, Song Y, Guo S - Med. Sci. Monit. (2015)

The analysis of recipient stromal cell replacement and gene expressions of recipient BMCs. (A) The gating protocol for donor-derived stromal cells in recipient BMCs. FSC, Forward Scatter. SSC, Side Scatter. Blue line, isotype control. (B, C) Donor stromal percentage in recipient BMCs were assessed at 1 (B) and 18 (C) weeks after BMT. Donor stromal cell population were H-2kb+ CD45-CD31-TER119- cells in BMCs harvested from the left tibia BMs. *, p value for IB-eBMCs versus each control group, p<0.05. Each dot represents an individual mouse result and bars are the mean for each group (%), n=6 per group. (D–G) Quantitative PCR analyzes of CXCL-12 (D, E) and n-cadherin (F, G) expression of recipient BMCs were performed at 1 week (D, F) and 18 weeks (E, G) after BMT. The corresponding gene expression level of BMCs from normal untreated mice was taken as control after normalization to GAPDH. All the bars represent the mean folds increase relative to that of normal mice BMCs for each group, n=6–8 per group. *, p value of IB-eBMCs group versus each control group (IB-cBMCs, IV-eBMCs, IV-cBMCs group respectively), p<0.05. ns is for not significant.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4588631&req=5

f1-medscimonit-21-2757: The analysis of recipient stromal cell replacement and gene expressions of recipient BMCs. (A) The gating protocol for donor-derived stromal cells in recipient BMCs. FSC, Forward Scatter. SSC, Side Scatter. Blue line, isotype control. (B, C) Donor stromal percentage in recipient BMCs were assessed at 1 (B) and 18 (C) weeks after BMT. Donor stromal cell population were H-2kb+ CD45-CD31-TER119- cells in BMCs harvested from the left tibia BMs. *, p value for IB-eBMCs versus each control group, p<0.05. Each dot represents an individual mouse result and bars are the mean for each group (%), n=6 per group. (D–G) Quantitative PCR analyzes of CXCL-12 (D, E) and n-cadherin (F, G) expression of recipient BMCs were performed at 1 week (D, F) and 18 weeks (E, G) after BMT. The corresponding gene expression level of BMCs from normal untreated mice was taken as control after normalization to GAPDH. All the bars represent the mean folds increase relative to that of normal mice BMCs for each group, n=6–8 per group. *, p value of IB-eBMCs group versus each control group (IB-cBMCs, IV-eBMCs, IV-cBMCs group respectively), p<0.05. ns is for not significant.
Mentions: All staining was performed at a cell concentration of 1×106/ml. All peripheral blood marrow cells (PBMCs) were gated according to the protocol described by Krause [31]. The peripheral blood chimerism level was calculated as H-2kb + cells percentage (%) in all PBMCs. The donor HSCs phenotype was H-2kb+ c-kit+ Sca-1+ Lineage- BMCs. Donor CFSE-positive cells were calculated as CFSE-positive cells in CD45+ cell population. The gate was set according to isotype control. The flow cytometry gating protocol for donor stromal cells in recipient BMCs is shown in Figure 1A. Donor stromal cell population are represented as H-2kb+CD45-CD31-TER119- cells in all BMCs (%).

Bottom Line: We compared the new method with each control group for allogeneic HSCs homing pattern, peripheral blood chimerism level, skin allograft survival time, and donor stromal cell percentage in recipient BM.By AMD3100 blockade at day 1, peripheral blood chimerism level and donor stromal cell percentage were significantly reduced as compared to the control group without AMD3100 blockade.Our study suggests that IBBMT of endosteal BMCs is an effective approach for HSCT in inducing allogeneic hematopoietic reconstitution.

View Article: PubMed Central - PubMed

Affiliation: Department of Plastic and Reconstructive Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, China (mainland).

ABSTRACT

Background: Hematopoietic stem cell transplantation (HSCT) has been considered as an effective approach at inducing allogeneic hematopoietic reconstitution and immune tolerance. However, it remains critical to find the optimal HSCT delivery method and robust sources of hematopoietic stem cells (HSCs).

Material and methods: We introduced a new method by infusing allogeneic endosteal bone marrow cells (BMCs) harvested from long bones endosteum through intra-bone marrow transplantation (IBBMT) into irradiated mice. Recipient mice that were transplanted with central BMCs or through intravenous bone marrow transplantation (IVBMT) were used as controls (n=6 per group). We compared the new method with each control group for allogeneic HSCs homing pattern, peripheral blood chimerism level, skin allograft survival time, and donor stromal cell percentage in recipient BM. AMD3100 was injected to determine whether chemokine stromal cell-derived factor-1 (CXCL-12) was critical for the new method.

Results: More allogeneic HSCs homed into spleen and bone marrow for the new method as compared to each control group. IBBMT of endosteal BMCs led to a higher peripheral blood chimerism and skin allograft survival. At 18 weeks, donor stromal cell percentage in recipient BMCs was higher for the new method than in each control group. By AMD3100 blockade at day 1, peripheral blood chimerism level and donor stromal cell percentage were significantly reduced as compared to the control group without AMD3100 blockade.

Conclusions: Our study suggests that IBBMT of endosteal BMCs is an effective approach for HSCT in inducing allogeneic hematopoietic reconstitution. The advantage is dependent upon the early expression of CXCL-12 after bone marrow transplantation.

No MeSH data available.


Related in: MedlinePlus